SUMMARY This renewal application tests several hypotheses H1 -The human gut microbiota is causally related to obesity and its associated metabolic abnormalities; H2 - The microbiota contains bacterial taxa that have effects on both adiposity and obesity-associated metabolic dysfunction including insulin-resistance, as well as taxa that exert selective effects on energy storage or metabolic activities disturbed in obese states; H3 - Diet influences expression of the activities of these groups of organisms; H4- The reduced diversity seen in the microbiota of obese individuals with and without metabolic dysfunction can be 'repaired' by adding microbes whose niches are not well-represented and dietary components that allow these microbes to establish themselves and express health-promoting functions. DK078669 will use a generally applicable translational research pipeline for microbiota-directed diagnostics and therapeutics [discovery of new probiotics, prebiotics, and synbiotics], Innovative features include (i) recruitment and detailed physiologic/metabolic phenotyping of same-sex female discordant twin-pairs, doubly or singly discordant for lean versus obese, and metabolic healthy versus unhealthy (insulin-resistant) states (abbreviated LnMH/ObMUN, LnMH/ObMH, ObMH/ObMUN), and implementation of controlled in-home diet studies (Project 2); (ii) transplantation of their intact uncultured gut communities, and subsequently extensive bacterial culture collections derived from their microbiota, into gnotobiotic mice fed the same NHANES-based diets as those consumed by the discordant twins during their in-home diet studies, to assess whether human gut communities can transmit their human donor phenotypes to recipient mice and the sensitivity of these transmitted phenotypes to diet (Project 1); (iii) co-housing mice, harboring transplanted microbiota from LnMH/ObMUN, LnMH/ObMH, ObMH/ObMUN pairs to identify taxa from LnMH (and then ObMH ) which ameliorate the increased adiposity and/or metabolic phenotypes associated with ObMUN microbiota, and to determine whether or not the effects are family-specific (to address the question of whether gut restoration has to be a within-family affair) (Project 1); (iv) development of new analytic tools for analyzing multi-omics time series studies of humans and mice (Project 3). Core A is an established metabolomics unit that will provide a combination of broad coverage and analytical precision for defining metabolic phenotypes in human subjects and derived gnotobiotic mouse models. Core B is a data repository for multi-omics datasets and their subsequent deposition in public databases. Our team of highly interactive, interdisciplinary, basic and clinical translational scientists has published 57 PPG papers during the current funding period. Our results suggest that in some obese humans, the gut microbiota is shifted to a state that can sustain obesity and its associated metabolic abnormalities, and that filling empty niches in Ob microbiota with Ln-derived taxa requires an diet that allows these taxa to be established and express their heath-promoting functions.